Standardization of organoid culture in cancer research.

Establishing a valid in vitro model to represent tumor heterogeneity and biology is critical but challenging. Tumor organoids are self-assembled three-dimensional cell clusters which are of great significance for recapitulating the histopathological, genetic, and phenotypic characteristics of primary tissues. The organoid has emerged as an attractive in vitro platform for tumor biology research and high-throughput drug screening in cancer medicine. Organoids offer unique advantages over cell lines and patient-derived xenograft models, but there are no standardized methods to guide the culture of organoids, leading to confusion in organoid studies that may affect accurate judgments of tumor biology. This review summarizes the shortcomings of current organoid culture methods, presents the latest research findings on organoid standardization, and proposes an outlook for organoid modeling.

A risk model based on pyroptosis subtypes predicts tumor immune microenvironment and guides chemotherapy and immunotherapy in bladder cancer.

Although immunotherapy has revolutionized bladder cancer (BLCA) therapy, only few patients demonstrate durable clinical benefits due to the heterogeneity. Emerging evidence has linked pyroptosis to shaping tumor microenvironment (TME) and predicting therapy response. However, the relationship between pyroptosis and immunotherapy response in BLCA remains elusive. In this study, we performed a comprehensive bioinformatic analysis to dissect the role of pyroptosis in BLCA. Differentially expressed pyroptosis-related genes (DEPRGs) between tumor and normal tissues were identified using publicly available datasets. Kaplan-Meier analysis was performed to screen for DEPRGs associated with survival. Consensus clustering was used for BLCA subtyping. TME characteristics were evaluated by CIBERSORT, ESTIMATE and immune checkpoint genes (ICGs). Following univariate COX regression and LASSO analyses with pyroptosis-related DEGs, the risk model and nomogram were constructed with TCGA dataset and validated in the GEO dataset. Furthermore, therapeutic responses in high- and low-risk groups were compared using TIDE and GDSC databases. Two pyroptosis-related subtypes (Cluster 1 and 2) were identified based on expression patterns of GSDMA and CHMP4C. Bioinformatic analyses showed that cluster 1 had poor survival, more M0/M1/M2 macrophages, higher immune/stromal/ESTIMATE scores, and higher expression levels of ICGs. A 15-gene signature for predicting prognosis could classify patients into high- and low-risk groups. Furthermore, the correlation of risk scores with TIDE score and IC50 showed that patients in low-risk group were more sensitive to immunotherapy, whereas patients in high-risk group could better benefit from chemotherapy. Our study identified two novel pyroptosis-related subtypes and constructed a risk model, which can predict the prognosis, improve our understanding the role of PRGs in BLCA, and guide chemotherapy and immunotherapy.

Propofol Disrupts Clear Cell Renal Cell Carcinoma Tumorigenesis by Regulating circFBXW7/miR-942 Axis.

BACKGROUND: Propofol is a commonly used intravenous anesthetic and has been found to perform anticancer effects in many cancers. However, the effects and mechanisms of propofol in clear cell renal cell carcinoma (ccRCC) remain largely undefined. METHODS: The expression of circular RNA FBXW7 (circFBXW7) and miR-942 was detected by qRT-PCR. Cell proliferation, apoptosis, migration, and invasion capacities were analyzed using cell counting kit-8, colony formation, flow cytometry, and transwell assays, respectively. Western blot was used to detect the expression levels of PCNA, Cleaved-caspase 3 and MMP protein. The bindings between miR-942 and circFBXW7 were verified using RNA pull-down, dual-luciferase reporter, and RIP assays. Xenograft tumor analysis was employed to detect tumorigenesis in vivo. RESULTS: Propofol alleviated cell proliferation, migration, invasion, and induced apoptosis in vitro and impeded tumor growth in vivo in ccRCC. Propofol elevated the level of circFBXW7, which knockdown reversed the anticancer effects of propofol on ccRCC cell tumorigenesis. CircFBXW7 directly bound to miR-942, and suppressed ccRCC cell malignant biological behaviors via targeting miR-942. Besides that, propofol decreased miR-942 expression, and miR-942 overexpression attenuated the effects of propofol on ccRCC cells. Moreover, propofol could regulate miR-942 expression through circFBXW7. CONCLUSION: Propofol suppressed the growth, migration, and invasion of ccRCC cells by regulating circFBXW7/miR-942 axis, suggesting a potential therapeutic strategy for the intervention of human ccRCC development.

Electro-characteristics of Myocardial Pouches and Reduction of the Frequency of Steam Pops During Radiofrequency Ablation.

Background: Radiofrequency ablation (RFA) effectively treats arrhythmia. Steam pop (SP) is a dangerous complication of RFA, which can lead to pericardial tamponade or even death. Objective: This study aimed to explore the electro-characteristics of myocardial pouches, and the relationship between SP, pouch, and impedance. Methods: Swine myocardium was divided into the pouch group and smooth myocardium group. Continuous RFA at 50 W was applied. The initial impedance reduction within the first 3 s of ablation and the time from the start of ablation to SP were recorded. After enabling the delta impedance cutoff function, RFA was performed at different percentage of delta impedance (PDI) cutoff thresholds. Results: The impedance was higher for the pouch myocardium compared to the smooth myocardium (123.22 ± 8.63 Ω and 95.75 ± 4.75 Ω, respectively; p < 0.001). The RFA duration before SPs was shorter in the pouch group compared to the smooth myocardium group [9 s (interquartile range, IQR: 6.25-13 s) and 33 s (IQR: 26.25-40.75 s), respectively; p < 0.001]. Within the first 3 s of RFA, impedance reduction (24.65 ± 6.57 Ω and 12.78 ± 3.35 Ω, respectively; p < 0.001) and PDI [19.18% (IQR: 16.39-24.20%) and 12.96% (IQR: 11.17-14.39%), respectively; p < 0.001] were greater in the pouch group compared to the smooth myocardium group. A PDI of 15% and delta time of 3 s effectively reduced the frequency of SPs without seriously affecting RFA use. Conclusion: SPs occur more frequently in the pouch area during RFA. Appropriate delta impedance cutoff settings (PDI: 15%; delta time: 3 s) can reduce the frequency of SPs and improve the RFA safety.

The Predictive Value of Neutrophil-to-Lymphocyte Ratio and Platelet-to-Lymphocyte Ratio Levels of Diabetic Peripheral Neuropathy.

OBJECTIVE: This study was designed to assess the levels of neutrophil-to-lymphocyte ratio (NLR) and platelet-to-lymphocyte ratio (PLR) in diabetes patients to determine their prognostic value in predicting the disease of diabetic peripheral neuropathy (DPN). METHODS: We recruited 225 diabetes cases from the department of endocrinology of Anhui Provincial Hospital from August 2018 to October 2019. A total of 103 patients without diabetic peripheral neuropathy (DPN) were followed up for 18 months, and the number of patients of newly diagnosed DPN was counted. According to the results of neuroelectrophysiological examination, these patients were divided into the diabetes mellitus (DM) without DPN group and the DM with DPN group. The general information and results of blood samples were collected. The collected data were compared between groups, and the receiver operating characteristic curve (ROC) was drawn. The follow-up data were compared between groups and Binary Logistic regression analysis was performed. RESULTS: Patients with DPN shared distinct characteristics. For example, the patients were older, and had higher levels of inflammatory indicators (ie, levels of PLR and NLR), and lower level of indirect bilirubin, compared with patients without DPN. According to the receiver operating characteristic curve analysis, for type 1 diabetes, PLR showed the highest area under the curve (0.753). For type 2 diabetes, NLR showed the highest AUC of 0.602. For the follow-up results, patients with newly diagnosed DPN bad higher NLR level. CONCLUSION: If patients of type 1 and type 2 diabetes are combined with elevated level of PLR and NLR, respectively, they are more likely complicated with DPN. NLR and PLR could be used as predictors to help clinicians screening for DPN in different types of diabetes. For type 1 diabetes, if patients who were without DPN had higher NLR level, the risk of developing DPN in the future will be greatly increased.

Polysaccharide-Based Lotus Seedpod Surface-Like Porous Microsphere as an Efficient Drug Carrier for Cancer Treatment.

BACKGROUND: This study aimed to evaluate the properties and functions of polysaccharide-based porous microsphere (PPM) for drug delivery, as well as its inhibitory effect on malignant tumors. MATERIALS AND METHODS: PPM was prepared using the inverse emulsion polymerization method. FT-IR measurements were conducted to measure the wavenumber of PPM. Particle size distribution was tested with a particle analyzer, and surface morphologies of PPM were observed using a scanning electron microscope (SEM). Dialysis method, Cell Counting Kit-8 (CCK-8), and cell apoptosis analysis were adopted to evaluate the drug release, cytotoxicity and biocompatibility of mitomycin-C (MMC), respectively. Finally, an in vivo study was performed in C57BL/6 mice to confirm the function of MMC-loaded PPM on tumor growth. RESULTS: FT-IR spectra proved the successful preparation of MMC-loaded PPM. PPM had an average size of 25.90 ± 0.34 µm and then increased to 30.10 ± 0.20 µm after drug loading. Under SEM, the surface morphology was lotus seedpod surface-like, with macropits on the surface and micropores in macropits. Compared with the free MMC group, MMC-loaded PPM exhibited a delayed drug release rate in a pH-dependent manner and higher cell viability. Flow cytometry results showed that the cell apoptosis in the PPM/MMC group was lower than that in the free MMC group. In vivo experiment revealed the inhibitory efficacy of MMC-loaded PPM on malignant tumors. CONCLUSION: In summary, MMC-loaded PPM exhibited favorable surface morphology, sustained drug release ability, nontoxicity and excellent biocompatibility, suggesting that PPM might be a potential drug carrier for tumor treatment.

P2X7 receptor antagonist BBG inhibits endoplasmic reticulum stress and pyroptosis to alleviate postherpetic neuralgia.

Postherpetic neuralgia (PHN) is the most common complication of acute herpes zoster. The treatment of PHN remains a challenge for clinical pain management. The present study investigated the P2X7 receptor antagonist brilliant blue G (BBG) whether inhibits endoplasmic reticulum stress and pyroptosis (a necrotic form of cell death) and alleviates PHN. Varicella zoster virus (VZV)-infected CV-1 cells were used to induce PHN model. Mechanical paw withdrawal thresholds were measured using an ascending series of von Frey filaments. Immunohistochemistry was used to detect the expression of P2X7R in nerve tissues. Western blot was used to determine the expression of endoplasmic reticulum (ER) stress and pyroptosis-related molecules. The expression of IL-1ß and IL-18 in tissue homogenate was detected by ELISA. The PHN rat has the lower paw withdrawal threshold, but higher expression of P2X7 in nerve tissues. And, endoplasmic reticulum stress was activated and pyroptosis was increased in PHN rats. BBG can decrease pain thresholds and reduce ER stress and pyroptosis in PHN rats. In addition, ER stress activator tunicamycin (TM) can reverse the effect of BBG on the paw withdrawal thresholds, endoplasmic reticulum stress, and pyroptosis. Therefore, P2X7 receptor antagonist BBG alleviates PHN by activating ER stress and reducing pyroptosis.

Structural Evolution from Noninterpenetrated to Interpenetrated Thorium-Organic Frameworks Exhibiting High Propyne Storage.

Two thorium-organic frameworks of [Th6O4(OH)4(TFBPDC)6(H2O)6]n (Th-TFBPDC) and [Th6O4(OH)4(TFBPDC)4(HCOO)4(H2O)6]n (Th-TFBPDC-i) constructed from the 3,3',5,5'-tetrakis(fluoro)biphenyl-4,4'-dicarboxylate (TFBPDC2-) ligand were obtained in a reaction. At an early stage of the reaction, the formation of the three-dimensional (3D) framework of Th-TFBPDC was discovered. At a later stage of the reaction, the complete product of Th-TFBPDC-i was obtained. The structural evolution from a noninterpenetrated network of Th-TFBPDC to a 2-fold interpenetrated network of Th-TFBPDC-i is a dissolution-recrystallization process and rationalized as the four equatorial TFBPDC2- ligands in an octahedral [Th6O4(OH)4(TFBPDC)12] unit were displaced by four formate ligands to form a [Th6O4(OH)4(TFBPDC)8(HCOO)4] unit via a ligand substitution reaction. The large pore volume as well as the strong interactions between the host framework and guest propyne (C3H4) molecules demonstrated by computational results endow the highly water-stable Th-TFBPDC with the best-performing C3H4 storage under ambient conditions. This work presents a rare example of structural evolution from a 3D noninterpenetrated network to a 2-fold 3D interpenetrated network and a highly promising metal-organic framework (MOF) for C3H4 storage with a C3H4 uptake of 8.16 mmol g-1 at 298 K.

Effects of Sulcus Vocalis Depth on Phonation in Three-Dimensional Fluid-Structure Interaction Laryngeal Models.

Sulcus vocalis is an indentation parallel to the edge of vocal fold, which may extend into the cover and ligament layer of the vocal fold or deeper. The effects of sulcus vocalis depth d on phonation and the vocal cord vibrations are investigated in this study. The three-dimensional laryngeal models were established for healthy vocal folds (0 mm) and different types of sulcus vocalis with the typical depth of 1 mm, 2 mm, and 3 mm. These models with fluid-structure interaction (FSI) are computed numerically by sequential coupling method, which includes an immersed boundary method (IBM) for modelling the glottal airflow, a finite-element method (FEM) for modelling vocal fold tissue. The results show that a deeper sulcus vocalis in the cover layer decreases the vibrating frequency of vocal folds and expands the prephonatory glottal half-width which increases the phonation threshold pressure. The larger sulcus vocalis depth makes vocal folds difficult to vibrate and phonate. The effects of sulcus vocalis depth suggest that the feature such as phonation threshold pressure could assist in the detection of healthy vocal folds and different types of sulcus vocalis.

Dysregulated Expression Levels of Circular RNAs Serve as Prognostic and Clinicopathological Markers in Gastric Cancer: a Meta-Analysis.

BACKGROUND: Circular RNAs (circRNAs) are novel biomarkers that are widely investigated in various cancers. There is increasing evidence that the expression levels of circRNAs are upregulated or downregulated in various cancers, but the overall prognostic efficiency of circRNAs in gastric cancer (GC) remains unclear. Therefore, this meta-analysis studies the relationship between circRNA expression and the prognosis of gastric malignancies. METHODS: A systematic search was conducted in the PubMed, Web of Science (WOS), EMBASE, and Cochrane Library databases. Eligible studies reporting on the associations of circRNAs with the clinicopathological characteristics and prognosis of GC patients were included. Pooled odds ratios (ORs) and 95% confidence intervals (CIs) were utilized to assess clinicopathological parameters. Hazard ratios (HRs) and 95% CIs were used to evaluate the prognostic value of circRNAs using RevMan 5.3 and Stata 15.1 software. RESULTS: Fifteen eligible studies, including 13 for clinicopathological features and 15 for prognosis, were included in our study. For clinicopathological parameters, the high expression of oncogenic circRNAs was significantly associated with poor clinicopathological features, and the high expression of tumor-suppressor circRNAs was associ-ated with better clinicopathological features. In terms of prognostic value, oncogenic circRNAs had a negative influence on overall survival (OS: HR = 2.27, 95% CI: 1.93 - 2.69), and the high expression of tumor suppressor circRNAs was related to improved survival outcomes (OS: HR = 0.56, 95% Cl: 0.44 - 0.72). CONCLUSIONS: This meta-analysis revealed that the expression of circRNAs might be a useful prognostic biomarker in gastric cancer.

Palladium-catalyzed selective B(3)-H arylation of o-carboranes with arylboronic acids at room temperature.

A palladium-catalyzed selective B(3)-H arylation of o-carboranes at room temperature has been developed using readily available arylboronic acids as the aryl source, and the corresponding 3-aryl-o-carboranes were obtained in good to excellent yields. This method provides a powerful synthetic route for constructing polysubstituted o-carborane derivatives.

The predictive value of serum amyloid A and C-reactive protein levels for the severity of coronavirus disease 2019.

This study was designed to assess the levels of human serum amyloid A (SAA) and C-reactive protein (CRP) in patients with coronavirus disease 2019 (COVID-19) to determine their prognostic value in predicting the severity of disease. Patients with COVID-19 who presented with acute respiratory distress syndrome (ARDS) shared distinct characteristics. For example, the patients were older, and had higher levels of inflammatory indicators [i.e., levels of CRP, SAA, procalcitonin (PCT), and interleukin-6; CRP-to-PCT ratio; SAA-to-CRP ratio; and neutrophil-to-lymphocyte ratio (NLR)], higher inflammatory cell counts (i.e., white blood cell count and neutrophil count), and lower lymphocyte counts compared with patients without ARDS. Patients without ARDS still exhibited mild illness and had elevated SAA levels but not CRP levels. In patients with elevated SAA and CRP levels, the NLR was statistically associated with disease severity. According to the receiver operating characteristic curve analysis, the combined predictive probability of CRP and SAA levels, along with white blood cell count, showed the highest area under the curve (AUC; 0.878), and was able to distinguish between patients with and without ARDS. The cut-off level for SAA to predict the severity of COVID-19 was 92.900, with a sensitivity of 95.8%, a specificity of 53.7%, and an AUC of 0.712. For patients with elevated levels of SAA but not CRP, a mild condition was predicted. For patients with elevated levels of both SAA and CRP, and a high NLR, a severe infection was predicted, requiring medical attention. Therefore, CRP and SAA levels demonstrate a prognostic value for predicting the severity of COVID-19.

Neuroprotective Effects of Dexmedetomidine Preconditioning on Oxygen-glucose Deprivation-reoxygenation Injury in PC12 Cells via Regulation of Ca2+-STIM1/Orai1 Signaling.

Dexmedetomidine (DEX), a potent and highly selective agonist for α2-adrenergic receptors (α2AR), exerts neuroprotective effects by reducing apoptosis through decreased neuronal Ca2+ influx. However, the exact action mechanism of DEX and its effects on oxygen-glucose deprivation-reoxygenation (OGD/R) injury in vitro are unknown. We demonstrate that DEX pretreatment reduced OGD/R injury in PC12 cells, as evidenced by decreased oxidative stress, autophagy, and neuronal apoptosis. Specifically, DEX pretreatment decreased the expression levels of stromal interaction molecule 1 (STIM1) and calcium release-activated calcium channel protein 1 (Orai1), and reduced the concentration of intracellular calcium pools. In addition, variations in cytosolic calcium concentration altered apoptosis rate of PC12 cells after exposure to hypoxic conditions, which were modulated through STIM1/Orai1 signaling. Moreover, DEX pretreatment decreased the expression levels of Beclin-1 and microtubule-associated protein 1A/1B-light chain 3 (LC3), hallmark markers of autophagy, and the formation of autophagosomes. In conclusion, these results suggested that DEX exerts neuroprotective effects against oxidative stress, autophagy, and neuronal apoptosis after OGD/R injury via modulation of Ca2+-STIM1/Orai1 signaling. Our results offer insights into the molecular mechanisms of DEX in protecting against neuronal ischemia-reperfusion injury.

Neuroprotective effects of 1-O-hexyl-2,3,5-trimethylhydroquinone on ischaemia/reperfusion-induced neuronal injury by activating the Nrf2/HO-1 pathway.

1-O-Hexyl-2,3,5-trimethylhydroquinone (HTHQ), a lipophilic phenolic agent, has an antioxidant activity and reactive oxygen species (ROS) scavenging property. However, the role of HTHQ on cerebral ischaemic/reperfusion (I/R) injury and the underlying mechanisms remain poorly understood. In the present study, we demonstrated that HTHQ treatment ameliorated cerebral I/R injury in vivo, as demonstrated by the decreased infarct volume ration, neurological deficits, oxidative stress and neuronal apoptosis. HTHQ treatment increased the levels of nuclear factor erythroid 2-related factor 2 (Nrf2) and its downstream antioxidant protein, haeme oxygenase-1 (HO-1). In addition, HTHQ treatment decreases oxidative stress and neuronal apoptosis of PC12 cells following hypoxia and reperfusion (H/R) in vitro. Moreover, we provided evidence that PC12 cells were more vulnerable to H/R-induced oxidative stress after si-Nrf2 transfection, and the HTHQ-mediated protection was lost in PC12 cells transfected with siNrf2. In conclusion, these results suggested that HTHQ possesses neuroprotective effects against oxidative stress and apoptosis after cerebral I/R injury via activation of the Nrf2/HO-1 pathway.

Disruption of CXCR6 Ameliorates Kidney Inflammation and Fibrosis in Deoxycorticosterone Acetate/Salt Hypertension.

Circulating cells have a pathogenic role in the development of hypertensive nephropathy. However, how these cells infiltrate into the kidney are not fully elucidated. In this study, we investigated the role of CXCR6 in deoxycorticosterone acetate (DOCA)/salt-induced inflammation and fibrosis of the kidney. Following uninephrectomy, wild-type and CXCR6 knockout mice were treated with DOCA/salt for 3 weeks. Blood pressure was similar between wild-type and CXCR6 knockout mice at baseline and after treatment with DOCA/salt. Wild-type mice develop significant kidney injury, proteinuria, and kidney fibrosis after three weeks of DOCA/salt treatment. CXCR6 deficiency ameliorated kidney injury, proteinuria, and kidney fibrosis following treatment with DOCA/salt. Moreover, CXCR6 deficiency inhibited accumulation of bone marrow-derived fibroblasts and myofibroblasts in the kidney following treatment with DOCA/salt. Furthermore, CXCR6 deficiency markedly reduced the number of macrophages and T cells in the kidney after DOCA/salt treatment. In summary, our results identify a critical role of CXCR6 in the development of inflammation and fibrosis of the kidney in salt-sensitive hypertension.

Mechanism of epithelial­mesenchymal transition inhibited by miR­203 in non­small cell lung cancer.

The aim of the present study was to investigate whether miR­203 can inhibit transforming growth factor­ß (TGF­ß)­induced epithelial­mesenchymal transition (EMT), and the migration and invasion ability of non­small cell lung cancer (NSCLC) cells by targeting SMAD3. In the present study, the expression levels of miR­203, SMAD3 mRNA and protein in NSCLC tissues were examined, as well as their corresponding paracancerous samples. The miR­203 mimics and miR­203 inhibitor were transfected into the H226 cell line. RT­qPCR was used to assess the expression levels of E­cadherin, Snail, N­cadherin and vimentin mRNA, and western blotting was performed to detect the expression levels of p­SMAD2, SMAD2, p­SMAD3, SMAD3 and SMAD4. The cell migration and invasion abilities were detected by Transwell assays. The target site of SMAD3 was predicted by the combined action between miR­203 and dual luciferase. The results revealed that the RNA levels of miR­203, compared with paracancerous tissues, were decreased in NSCLC tissues, while SMAD3 mRNA and protein levels were upregulated, and miR­203 inhibited SMAD3 expression. Induction of TGF­ß led to decreased E­cadherin mRNA levels, upregulation of Snail, N­cadherin and vimentin mRNA levels (P<0.05), and significant increase in cell migration and invasion, whereas transfection of miR­203 mimics reversed the aforementioned results (P<0.05). Conversely, miR­203 inhibitor could further aggravate the aforementioned results (P<0.05). Western blot results revealed that transfection of miR­203 mimics significantly reduced the protein expression of SMAD3 and p­SMAD3 (P<0.05). Furthermore, the results of the Dual­Luciferase assay revealed that miR­203 inhibited SMAD3 expression by interacting with specific regions of its 3'­UTR. Overall, a novel mechanism is revealed, in which, miR­203 can inhibit SMAD3 by interacting with specific regions of the 3'­UTR of SMAD3, thereby restraining TGF­ß­induced EMT progression and migration and invasion of NSCLC cells.

Medi-Care AI: Predicting medications from billing codes via robust recurrent neural networks.

In this paper, we present an effective deep prediction framework based on robust recurrent neural networks (RNNs) to predict the likely therapeutic classes of medications a patient is taking, given a sequence of diagnostic billing codes in their record. Accurately capturing the list of medications currently taken by a given patient is extremely challenging due to undefined errors and omissions. We present a general robust framework that explicitly models the possible contamination through overtime decay mechanism on the input billing codes and noise injection into the recurrent hidden states, respectively. By doing this, billing codes are reformulated into its temporal patterns with decay rates on each medical variable, and the hidden states of RNNs are regularized by random noises which serve as dropout to improved RNNs robustness towards data variability in terms of missing values and multiple errors. The proposed method is extensively evaluated on real health care data to demonstrate its effectiveness in suggesting medication orders from contaminated values.

Danzhi Jiangtang Capsule Mediates NIT-1 Insulinoma Cell Proliferation and Apoptosis by GLP-1/Akt Signaling Pathway.

OBJECTIVE: This study aimed to investigate the effects of Danzhi Jiangtang Capsule (DJC) on the proliferation and apoptosis functions of NIT-1 pancreatic ß-cells exposed to high-glucose load through GLP-1 activated Akt/ FoxO1 signaling pathway. METHODS: Cellular apoptosis of NIT-1 pancreatic ß-cells was induced by culturing in medium with 33.3mmol/L high glucose (HG). Then low-dose DJC (HG +LD), high-dose DJC (HG +HD), high-dose DJC+ GLP-1 inhibition (HG +HD +GI), and high-dose DJC+AKT inhibition (HG +HD+AI) were added, respectively. Cellular proliferation was accessed by cell counting kit (CCK-8) and cellular apoptosis was measured by Annexin V-FITC/PI staining. The protein levels of phosphorylated phosphatidylinositol-3-kinase (p-PI3K), phosphorylated AKT (p-AKT), phosphorylated Forkhead box protein O1 (p-FoxO1), and cleaved caspase-3 were detected by Western blotting. The mRNA expression of pancreatic duodenal homeobox-1 (PDX-1), CyclinD1, Bcl-2, and insulin was tested by Q-PCR. RESULTS: Comparing to HG group, (HG+HD) group showed a significantly increased cellular proliferation. The apoptosis of NIT-1 cells also was obviously reduced, with downregulated cleaved caspase-3 protein level and upregulated PDX-1, CyclinD1, and Bcl-2 mRNA levels (P<0.05). Additionally, (HG+HD) group manifested increased insulin mRNA expression; the protein levels of p-PI3K and p-AKT were markedly increased and p-FoxO1 was decreased. All of the above therapeutic effects by DJC intervention had been reversed by GLP-1 inhibition in (HG+HD+GI) group or AKT inhibition in (HG+HD+AI) group. CONCLUSION: DJC was able to attenuate the toxicity of high-glucose load in NIT-1 pancreatic ß-cells, ascribed to the improvement of cellular proliferation and apoptosis by GLP-1/Akt signaling pathway. This study could supply a new mechanism of DJC effects on type 2 diabetes mellitus (T2DM) treatment.

Mitochondrial dysfunction in neurodegenerative diseases and drug targets via apoptotic signaling.

Mitochondrial dysfunction is becoming one of the most emerging pathological process in the etiology of neurological disorders. Other common etiologies of the neurological disorders are aging and oxidative stress. Neurodegenerative disorders for instance Huntington's disease, Parkinson's disease, Amyotrophic lateral sclerosis, Epilepsy, Schizophrenia, Multiple sclerosis, Neuropathic pain and Alzheimer's disease involves mitochondrial dysfunction and is regarded as the core of their pathological processes. Most central pathological feature of the neurodegenerative diseases is apoptosis which is regulated by mitochondria. Altered signaling of the apoptotic mechanisms are involved in neurodegeneration. Abnormal levels of these molecular apoptotic proteins promotes the pathogenesis of neurological disorders. Mitochondria are also implicated in the production of reactive oxygen species (ROS). Raised ROS levels initiates the cascade leading to the non-apoptotic death of cells. ROS produced in cells acts as signaling molecules, but when produced in abundance will result in cellular consequences to deoxyribonucleic acid, proteins and lipids, decreased effectiveness of cellular mechanisms, initiation of inflammatory pathways, excitotoxicity, protein agglomeration and apoptosis. Protecting mitochondrial function has been identified as the most effective therapeutic approach to attenuate the pathogenesis of neurodegenerative diseases. This review aims to provide an insight into the mitochondrial dysfunction in the pathogenesis of neurological disorders, alteration in signaling cascades of apoptosis in mitochondrial dysfunction and the therapeutic strategies (both natural and synthetic drugs) targeting these mitochondrial apoptotic pathways and oxidative stress that holds great promise.

Gephyrin suppresses lung squamous cell carcinoma development by reducing mTOR pathway activation.

Background: The mTOR pathway is altered in a multitude of cancers, including lung cancer; however, abnormal activation in this pathway is less common in lung adenocarcinoma (LUAD) than in lung squamous cell carcinoma (LUSC). Gephyrin is a highly conserved and widely expressed ancient protein in vertebrate tissues. Its role and molecular mechanism in lung cancer development are largely unknown. Method: We analyzed the expression profile of gephyrin and overall survival rates in LUAD and LUSC. The LUSC cells (H520 and SK-MES-1) were transfected with pLV-gephyrin to establish gephyrin stable overexpression cell lines. Real-time quantitative PCR and Western blot were performed to detect the mRNA and protein levels. The cell growth and cell cycle were detected by the MTT assay and flow cytometry. Finally, a xenograft tumor model was established to determine cell tumorigenesis in vivo. Results: Our results show that gephyrin was reduced in LUAD and LUSC, and its low expression in LUSC patients indicated poor prognosis. Gephyrin overexpression suppressed LUSC cell proliferation, arrested cell cycle progression, and decreased the expression of cell-cycle related proteins such as cyclin D1, cyclin-dependent kinase-2 (CDK2), and proliferation-related protein proliferating cell nuclear antigen (PCNA). Conversely, knockdown of gephyrin promoted LUSC cell growth. Moreover, gephyrin reduced mTOR pathway activation to inhibit cyclin D1 and CDK2 translation. Mechanistically, gephyrin suppressed mTOR pathway activation by promoting mTOR degradation. Furthermore, gephyrin overexpression suppressed LUSC tumorigenesis. Conclusion: Gephyrin suppressed LUSC development by reducing mTOR pathway activation, implicating gephyrin as a potential molecular target for LUSC management.